1. Field of the Invention
The present invention relates to the preclinical testing of therapeutic products and, more particularly, to the testing of a protein, intended for repeated administration in humans, for immunogenicity through the use of a protocol involving the use of the immune system of an animal and the comparison of immune responses to a test and a reference protein.
2. Description of the Related Art
With the advent of recombinant DNA technology, a number of therapeutically active peptides have been developed for use in humans. These peptides are coded for by human genes which have been cloned into a host system for production. The host system may be a bacterium, such as E. coli, a yeast such as Saccharomyces cerevisae, or a mammalian cell line, such as a hybridoma or a continuous cell line such as Chinese Hamster Ovary or Baby Hamster Kidney.
Regardless of the host system chosen, there are questions which may be raised as to the "authenticity" of a peptide product, in terms of its suitability for human use. One such question involves the response of an immunocompetent human host to the therapeutic peptide. This response may in certain instances have clinical significance, such as has been reported in some cases of administration of recombinant human growth hormone (produced in E. coli) or in the case of murine monoclonal antibodies. Unfortunately, the human immune response to a therapeutic peptide is impossible to predict with certainty, and little literature exists on the development of animal models which can be used to predict immunogenicity in preclinical testing.
One method which has been previously used by developers of therapeutic peptides is to simply administer to animals repeated injections of the protein of interest and observe clinical signs. This method has several substantial drawbacks. First, it is expected that all peptides beyond approximately 5kD will elicit an immune response in a non-homologous species. Therefore, the appearance of antibodies in such a protocol is to be expected. The mere quantification of these antibodies is not particularly informative, since comparisons among different peptides and different animals are not meaningful.
A common method of immunogenicity testing in animals involves repeat administration of final container product and subsequent animal evaluation. Such evaluation may range from observation for anaphylactic reactions to measurement of immune complexes.
Another methodology of relevance is passive cutaneous anaphylaxis (PCA), although this test method is not particularly used for immunogenicity testing per se. This is because it is a passive system and does not measure immune response. In this system, an antibody is administered to a guinea pig intracutaneously. Then, an antigen of interest is administered intravenously, coupled with a blue dye.
If antigen-antibody complexes are formed, the complexes and the dye will be extravacised, leading to blue spot(s) at the injection site(s). This method is further described by Ovary, Z. (1958).
A chemotactic assay for immunogenicity is described in U.S. Pat. No. 4,714,674.